Defining the role of HMGA2 in driving tRNA synthesis in pancreatic ductal adenocarcinoma

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Pancreatic Ductal Adenocarcinoma (PDA) is an extremely lethal disease with a 5-year survival rate of less than 10% and is likely to become the second leading cause of cancer related deaths by 2030. We know that pancreatic cancer is complex given that patients respond very differently to chemotherapy. To understand these differences, researchers have recently characterized two main subtypes: classical/pancreatic progenitor and quasi-mesenchymal (QM)/basal/squamous. The QM PDA subtype is more aggressive and has the worst overall survival. Understanding what defines each subtype, their susceptibilities and mechanisms of resistance will help to identify new targeted therapies for this devastating disease. While investigating these questions the Kugel lab made the startling discovery that the QM PDA subtype expresses high levels of high-mobility group (HMG) protein A2 (HMGA2) and have evolved mechanisms to dramatically increase protein translation. This raised the intriguing possibility that the aberrant activation of HMGA2 increases protein translation and may be one mechanism by which PDA attains the highly aggressive QM phenotype. The aim of this proposal is to elucidate the role of HMGA2 in regulating tDNA transcription in QM PDA, determine the role of differentially expressed tRNAs on protein synthesis and finally, explore the role of HMGA2 in PDA subtype determination, initiation and maintenance. Therefore, the Cancer Genome Atlas will be utilized to investigate potential candidate tRNAs, that will be validated in HMGA2 (HMGA2high/QM PDA) cell lines against HMGA2low/classical PDA cell lines. The direct influence of HMGA2 on tRNA expression will be tested in classical or QM PDA cell lines engineered to either overexpress WT HMGA2 or silence HMGA2 by siRNA, respectively by tRNA sequencing. HMGA2 acts as architectural transcription factor. We will assay the effect of HMGA2 on chromatin structure to induce tDNA transcription by ChIP analysis of canonical and non-canonical histones bound to candidate tRNA gene bodies in engineered PDA cell lines. We will perform ribosome profiling to identify changes in the protein synthesis due to differentially expressed tRNAs. Lastly, the engineered cell lines will be used to evaluate the effect of HMGA2 on QM-markers like increased TP63DeltaN and mesenchymal markers expression, increased chromatin bound MYC levels and reduced sensitivity to genetic ablation of KRAS. Understanding the epigenetic switch and the need of aberrant protein synthesis in QM-PDA will allow us to better understand this highly aggressive PDA subtype. The findings of this proposal could be the base to investigate into new drugs and therapeutic plans to increase the therapeutic chances of this lethal disease.